人羊膜上皮细胞分化为结膜上皮细胞实验研究

Experimental study on the differentiation from human amniotic epithelial cells to conjunctival epithelium cells

目的探讨人羊膜上皮细胞在特定环境下诱导分化为结膜上皮细胞的可能性。方法人羊膜上皮细胞与去上皮结膜基质的结膜匀浆培养液孵育,角蛋白CK4、CK13、MUC5AC免疫组化鉴定细胞表型;过碘酸-Schiff试剂（PAS）;酶联免疫吸附（ELISA）法检测不同时间培养液内MUC5AC水平;逆转录-聚合酶链反应（RT-PCR）检测分化细胞MUC5AC基因表达。结果人羊膜上皮细胞经诱导培养后,免疫组化显示角蛋白CK4、CK13、MUC5AC表达阳性;PAS染色阳性;ELISA检测1~7 d内培养液中均有微量MUC5AC且逐渐升高;RT-PCR检测分化细胞MUC5AC表达阳性。结论人羊膜上皮细胞具有干细胞分化潜能,在特定条件下可定向分化为具有一定生理功能的结膜上皮细胞。

Objective To trans-differentiate from human amniotic epithelial cells to conjunctival epithelium under the specific condition.Methods Human amniotic epithelial cells were cultured under the inducing conditions of denuded conjunctival matrix and conjunctival homogenate.Immunohistochemistry with cytokeratin 4（CK4）,CK13,MUC5AC,and PAS staining were performed respectively.The levels of MUC5AC in different time were measured by ELISA.The gene expressions of intracellular MUC5AC were tested by RT-PCR.Results After the induction of human amniotic epithelial cells,the expression of CK4,CK13 and MUC5AC in the test of immunohistochemistry was positive,and the positive cells of purplish red color were detected in PAS staining.Trace amount of MU5AC in culture medium was measured by ELISA and it increased gradually in day 1-7.The expression of MUC5AC in differentiated cells was positive shown by RT-PCR.Conclusion Human amniotic epithelial cells have a potential to differentiate to stem cells.In a specific condition,they can differentiate to conjunctival epithelium cells.